A photoacoustic microscope system provides label-free multilayered histology-like imaging of unprocessed human breast specimens.

Three-dimensional (3D) optical imaging of whole biological organs with microscopic resolution has remained a challenge. Most versions of such imaging techniques require special preparation of the tissue specimen. Here we demonstrate microtomy-assisted photoacoustic microscopy (mPAM) of mouse brains and other organs, which automatically acquires serial distortion-free and registration-free images with endogenous absorption contrasts. Without tissue staining or clearing, mPAM generates micrometer-resolution 3D images of paraffin- or agarose-embedded whole organs with high fidelity, achieved by label-free simultaneous sensing of DNA/RNA, hemoglobins, and lipids. mPAM provides histology-like imaging of cell nuclei, blood vessels, axons, and other anatomical structures, enabling the application of histopathological interpretation at the organelle level to analyze a whole organ. Its deep tissue imaging capability leads to less sectioning, resulting in negligible sectioning artifact. mPAM offers a new way to better understand complex biological organs.

Acquiring ultrafast and high spectral resolution optical images is key to measure transient physical or chemical processes, such as photon propagation, plasma dynamics, and femtosecond chemical reactions. At a trillion Hz frame rate, most ultrafast imaging modalities can acquire only a limited number of frames. Here, we present a compressed ultrafast spectral-temporal (CUST) photographic technique, enabling both an ultrahigh frame rate of 3.85 trillion Hz and a large frame number. We demonstrate that CUST photography records 60 frames, enabling precisely recording light propagation, reflection, and self-focusing in nonlinear media over 30 ps. CUST photography has the potential to further increase the frame number beyond hundreds of frames. Using spectral-temporal coupling, CUST photography can record multiple frames with a subnanometer spectral resolution with a single laser exposure, enabling ultrafast spectral imaging. CUST photography with high frame rate, high spectral resolution, and high frame number in a single modality offer a new tool for observing many transient phenomena with high temporal complexity and high spectral precision.

Histological images can reveal rich cellular information of tissue sections, which are widely used by pathologists in disease diagnosis. However, the gold standard for histopathological examination is based on thin sections on slides, which involves inevitable time-consuming and labor-intensive tissue processing steps, hindering the possibility of intraoperative pathological assessment of the precious patient specimens. Here, by incorporating ultraviolet photoacoustic microscopy (UV-PAM) with deep learning, we show a rapid and label-free histological imaging method that can generate virtually stained histological images (termed Deep-PAM) for both thin sections and thick fresh tissue specimens. With the tissue non-destructive nature of UV-PAM, the imaged intact specimens can be reused for other ancillary tests. We demonstrated Deep-PAM on various tissue preparation protocols, including formalin-fixation and paraffin-embedding sections (7-µm thick) and frozen sections (7-µm thick) in traditional histology, and rapid assessment of intact fresh tissue (~ 2-mm thick, within 15 min for a tissue with a surface area of 5 mm × 5 mm). Deep-PAM potentially serves as a comprehensive histological imaging method that can be simultaneously applied in preoperative, intraoperative, and postoperative disease diagnosis.